Biomolecule immobilization method

Abstract

In an example method, a grafting solution is applied to a patterned substrate using a liquid-phase thin-film deposition technique. The patterned substrate includes a lane surrounded by, or a plurality of depressions separated by interstitial regions; and a polymer in the lane or in each of the plurality of depressions. The polymer is functionalized with a first click reaction moiety. The grafting solution includes a solvent; a polymer matrix material dissolved in the solvent; and primers of a primer set dissolved in the solvent, each of the primers being terminated with a second click reaction moiety. The applied grafting solution is dried. During drying, a solid polymer matrix is formed and at least some of the primers attach to the polymer i) via the first and second click reaction moieties and ii) in at least a portion of the lane or in at least some of the plurality of depressions.

What is claimed is: 
     
         1 . A method, comprising:
 using a liquid-phase thin-film deposition technique, applying a grafting solution to a patterned substrate, wherein:
 the patterned substrate includes:
 a lane surrounded by interstitial regions or a plurality of depressions separated by interstitial regions; and 
 a polymer in the lane or in each of the plurality of depressions, the polymer being functionalized with a first click reaction moiety; and 
 
 the grafting solution includes:
 a solvent; 
 a polymer matrix material dissolved in the solvent; and 
 primers of a primer set dissolved in the solvent, each of the primers being terminated with a second click reaction moiety; and 
 
   drying the grafting solution, thereby forming a solid polymer matrix and attaching at least some of the primers to the polymer i) via the first and second click reaction moieties and ii) in at least a portion of the lane or in at least some of the plurality of depressions.   
     
     
         2 . The method as defined in  claim 1 , further comprising removing the solid polymer matrix and unattached primers by rinsing the patterned substrate with a solvent of the solid polymer matrix, whereby the attached primers remain in the at least the portion of the lane or the at least some of the plurality of depressions. 
     
     
         3 . The method as defined in  claim 2 , further comprising:
 introducing a protective coating solution to the patterned substrate; and   drying the protective coating solution to form a solid protective coating.   
     
     
         4 . The method as defined in  claim 3 , wherein prior to introducing the protecting coating solution or after drying the protective coating solution, the method further comprises bonding the patterned substrate to a second patterned substrate or to a lid to form a flow cell. 
     
     
         5 . The method as defined in  claim 1 , wherein while the solid polymer matrix and unattached primers are present in the at least the portion of the lane or in the at least some of the plurality of depressions, the method further comprises bonding the patterned substrate to a second patterned substrate or to a lid to form a flow cell. 
     
     
         6 . The method as defined in  claim 1 , wherein the grafting solution includes:
 up to 15% (mass to volume) of the polymer matrix material; and   up to 10% (mass to volume) of the primers.   
     
     
         7 . The method as defined in  claim 1 , wherein the polymer matrix material is selected from the group consisting of a water-soluble non-cationic synthetic polymer; a water-soluble natural polysaccharide or a derivative thereof; a water-soluble natural protein or a derivative thereof; and combinations thereof. 
     
     
         8 . The method as defined in  claim 1 , wherein the grafting solution further includes an additive selected from the group consisting of a sugar, a surfactant, and combinations thereof. 
     
     
         9 . The method as defined in  claim 8 , wherein:
 the grafting solution includes:
 up to 15% (mass to volume) of the polymer matrix material; 
 up to 35% (mass to volume) of the additive; and 
 up to 10% (mass to volume) of the primers; and 
   a total (mass to volume) of the polymer matrix material and the additive ranges from about 5% to about 40%.   
     
     
         10 . The method as defined in  claim 9 , wherein:
 the polymer matrix material includes a water-soluble non-cationic synthetic polymer;   the water-soluble non-cationic synthetic polymer is a polyvinyl alcohol/polyethylene glycol graft copolymer;   the additive is the sugar; and   the sugar is sucrose.   
     
     
         11 . The method as defined in  claim 1 , wherein the liquid-phase thin-film deposition technique is selected from the group consisting of dispense coating, slot-die coating, and inkjet printing. 
     
     
         12 . The method as defined in  claim 1 , wherein the first click reaction moiety is azide and the second click reaction moiety is bicyclononyne. 
     
     
         13 . The method as defined in  claim 1 , wherein the first click reaction moiety is azide and the second click reaction moiety is dibenzocyclooctyne. 
     
     
         14 . The method as defined in  claim 1 , wherein the first click reaction moiety is tetrazine and the second click reaction moiety is trans-cyclooctene. 
     
     
         15 . The method as defined in  claim 1 , wherein drying the grafting solution is accomplished at ambient temperature. 
     
     
         16 . The method as defined in  claim 1 , wherein drying the grafting solution involves exposing the applied grafting solution to heat ranging from about 25° C. to about 120° C. 
     
     
         17 . The method as defined in  claim 1 , further comprising allowing the solid polymer matrix to remain in the lane or the plurality of depressions during shipping, storage, or combinations thereof. 
     
     
         18 . The method as defined in  claim 1 , wherein:
 the patterned substrate is part of a flow cell including:
 the patterned substrate bonded to a lid or a second patterned substrate; and 
 a flow channel defined between the patterned substrate and the lid or the second patterned substrate; 
   the liquid-phase thin-film deposition technique is a flow through technique; and   drying the grafting solution involves introducing an air or nitrogen gas bubble into the flow cell while the grafting solution is present in the flow channel.   
     
     
         19 . A method, comprising:
 using a liquid-phase thin-film deposition technique, applying a grafting solution to a patterned substrate, wherein:
 the patterned substrate includes:
 a lane surrounded by interstitial regions or a plurality of depressions separated by interstitial regions; and 
 a polymer in the lane or in each of the plurality of depressions, the polymer being functionalized with a first click reaction moiety; and 
 
 the grafting solution includes:
 a solvent; 
 a polymer matrix material dissolved in the solvent; and 
 a plurality of biomolecules dissolved in the solvent, each of the plurality of the biomolecules being terminated with a second click reaction moiety; and 
 
   drying the grafting solution, thereby forming a solid polymer matrix and attaching at least some of the plurality of biomolecules to the polymer i) via the first and second click reaction moieties and ii) in at least a portion of the lane or in at least some of the plurality of depressions.   
     
     
         20 . The method as defined in  claim 19 , wherein the plurality of biomolecules is selected from the group consisting of biotin, a protein, an enzyme, DNA, RNA, an aptamer, or a combination of a primer set and a quality control oligonucleotide.